The research proposed here is directed toward an understanding of the molecular mechanism of homologous genetic recombination. This goal is approached through the study of special sites in DNA promoting a high rate of recombination in their vicinity and through the identification and study of the activity of proteins interacting with these sites. The Chi recombinational hotspots of the bacterium Escherichia coil and the M26 recombinational hotspot of the fission yeast Schizosaccharomyces pombe will be studied in the proposed research. The interaction of Chi and RecBCD enzyme, which directly activates Chi, and the Mts1,2 protein, which binds to the M26 site, will be studied. The regional specificity of the S. pombe rec8, rec1O, and rec11 gene products, hypothesized to act at hotspots, will be studied. These studies will elucidate the molecular mechanisms by which recombinational hotspots act and thereby elucidate an important (rate-limiting) step in homologous recombination. Specific aims are to: (A) determine the role of an essential RNA subunit of RecBCD enzyme in its interaction with Chi, (B) assess the mode of RecBCD-Chi interaction in E. coli cells, (C) determine the physical alteration of RecBCD upon cutting at Chi, (D) determine which strand of Chi is recognized by RecBCD, (B)test a model for Chi-stimulated conjugational and transductional recombination, (F)clone the mts1,2 genes and study mutants lacking the Mts1,2 protein, (G) determine the basis for the chromosomal context-dependence of M26, and (H) study the regional specificity of the Rec8, 10, and 11 activators of meiotic recombination. Recombination plays an important role in generating diversity among individuals in a population and, in some cases, among cells within an organism. Understanding the mechanism of recombination thus lends insight into the evolution and development of organisms. Aberrations of recombination may be responsible for certain chromosomal rearrangements associated with birth defects, hereditary diseases, and cancer.